The long term objective of this research proposal is to understand the cellular and molecular events underlying memory and learning in the mammalian central nervous system (CNS). The cerebellum is a component of the CNS that serves an important role in the learning and memory of motor tasks. While the cellular and molecular details of cerebellar learning are not well understood, changes in the efficacy of synaptic transmission between granule cells and Purkinje cells (PCs) are thought to play a central role. These changes could be expressed as modifications in the membrane properties of PCs. Current evidence indicates that glutamate is the neurotransmitter released by granule cells, depolarizing the membrane potential of and increasing cyclic nucleotide synthesis in PCs. Electrophysiological analyses of glutamate receptor types and cyclic nucleotide actions in PCs in vivo have yielded results that are difficult to interpret due to the likely possibility of indirect actions. The immediate objective of this proposal is to study the effects of glutamate and cyclic nucleotides in cultured PCs where a more direct analysis should be possible. Preliminary studies indicate that (a) PCs can be identified in culture using cell-specific immunocytochemical staining, (b) PCs cultured from rat embryos develop electrophysiological properties similar to relatively mature PCs isolated from two-week old rats, and (c) glutamate depolarizes the membrane potential of cultured PCs. The specific aims of this research proposal are the following: (1) to characterize the types of voltage-dependent and glutamate- activated conductances in cultured PCs using whole-cell patch recording techniques: (2) to use immunocytochemical staining procedures for detecting cyclic nucleotide levels in cultured PCs, and for assessing whether neurotransmitters and pharmacological agents known to elevate cyclic nucleotide levels in PCs in vivo can do the same in culture; (3) to investigate the effects of cyclic nucleotide elevation on the excitability and membrane ionic conductance of cultured PCSs. It is expected that the results of these studies will advance our understanding of the role of glutamate and cyclic nucleotides in granule cell-PC transmission, and the results may suggest possible cellular and molecular mechanisms underlying synaptic plasticity in the mammalian CNS. In the broader context,k this research may offer new insight into the etiology of diseases associated with learning impairment and memory loss.